Purification and Characterization of a β-Galactosidase from Peach (Prunus persica)

Abstract

A β-galactosidase (EC 3.2.1.23) from peach (Prunus persica cv Mibackdo) was purified and characterized. The purified peach β-galactosidase was 42 kDa in molecular mass and showed high enzyme activity against a the β-galactosidase substrate, ρ-nitrophenyl-β-d-galactopyranoside. The Km and Vmax values of the enzyme activity of the peach β-galactosidase were 5.16 and 0.19 mM for ρ-nitrophenyl-β-d-galactopyranoside mM/h, respectively. The optimum pH of the enzyme activity was pH 3.0, but it was relatively stable from pH 3.0–10.0. The temperature optimum was 50°C. The enzyme activities were not improved in the buffers that contained Ca2+, Cu2+, Zn2+, and Mg2+, which indicates that the purified peach β-galactosidase did not require these cations as co-factors. However, the enzyme was completely inhibited by Hg2+. The purified protein was cross-reacted with an antibody against the persimmon fruit β-galactosidase. A further comparison of the N-terminal amino acid sequence of the purified protein showed high homologies to those of β-galactosidase in apple (87%), persimmon (80%), and tomato (87%). Therefore, enzymatic, immunological, and molecular evidences in this study indicate that the purified 42-kDa protein is a peach β-galactosidase.